I am doing experiments on wheat leaves to isolate mitochondrial proteins. I have seen many references stating that we don't add BSA (in grinding and washing buffer) for isolation of proteins if we have to do western blotting in future. First of all, is it true? Secondly, what is the reason behind not using BSA for western blotting.
The BSA was served as an indicator to make sure all the isolation processes is equally in the volume or concentration. Usually, the BSA was used in blue native PAGE. If you are going to do the western blotting for desired protein, you can use Ponceus S red staining as loading control instead of BSA.
Hi,
Sometimes BSA is used as negative control on western blotting when cross-reactivity with antibodies but due to non-specific binding,its being avoided.I hope will help you.
All the best!
- During western blotting BSA is generally used as a blocking reagent ( to prevent non speific binding of antibodies)
- If the concentration rises much above 5% however it can start to mask specific epitopes bound to your membrane and thus will reduce your signal
- This is the reason that protein solutions with BSA much above 5% are not recommended
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